Eastern equine encephalomyelitis virus (EEEV) is a serious infection of humans, and is listed as a high- consequence select agent. In studies carried out during the previous funding period, we demonstrated that EEEV ecology in the mid- Southeastern USA differs fundamentally from that in the North Atlantic states. We have shown that ornithophilic vectors for EEEV exhibit temporally shifting preferences for avian hosts, a finding that suggests that these vectors preferentially target nestlings and young-of-the-year (YOY). We have developed a model that suggests that feeding on nestlings and YOY is an important factor driving the transmission of EEEV. We have also found evidence for EEE infection very early in the year and in species that feed primarily upon reptiles and amphibians. The overall goal of this project will be to build upon these findings to further understand how the interaction of EEEV, its vectors and vertebrate hosts results in maintenance of the virus, the development of avian enzootics and the escape of the virus from enzootic foci to infect humans and horses. The specific aims of the project are: 1. To test the hypothesis that EEEV vector mosquitoes feed preferentially upon YOY, using a microsatellite- based method to identify avian blood meals to the individual level. 2. To estimate the foraging range of Cx. erraticus, the major EEEV vector in the Southeast. 3. To measure EEEV exposure and reservoir competence of amphibians and reptiles. 4. To determine the vector competence of Cx. erraticus, Cx. peccator and Ur. sapphirina for EEEV. 5. To test the hypothesis that certain over-wintering mosquitoes are involved in maintaining the virus during the winter months. Together, these efforts will allow us to identify the most significant vectors for the virus, the potential mechanism for over-wintering, non-avian reservoirs, and the dynamics of spread of the virus from its enzootic focus to surrounding areas. When parameterized in our model, these data will allow us to predict the most important factors responsible for the development of an avian epizootic and for the spread of that epizootic to other species, including humans.